Name: the application of permanent middle cerebral artery ligation in the mouse
Uploaded: 2011-07-25T13:00:00
Description: Watch this Scientific Journal Video about The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse at JoVE.com

The surgical technique was originally acquired in the lab of Dr. William D. Hill at the Medical College of Georgia. The authors would also like to thank Dr. David A. Rempe and Landa Prifti for the use of the dissection camera. This research was supported by NIH NS041744, NS051279, F31 NS064700 and AHA 30815697D.

This protocol was approved by the University of Rochester committee devoted to the ethical use of animals in research (UCAR). Aseptic techniques should be followed during the protocol. The use of sterile gloves and a mask is required.
All the equipment, materials, chemicals, and tools that are used during the protocol are described in Table 1. 
1. Pre-surgical Preparation
<ol>
<li>Inject mice subcutaneously with buprenorphine (0.05mg/kg) 2 h before surgery, immedi...

<ol>
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The permanent MCA ligation method gives highly reproducible infarct volumes and increased post-operative survival rates compared to other methods available. The ease and the short duration (&tilde;30 min) of the procedure make it even more practical. The method is widely used in both mice and rats.
This technique requires an invasive surgery under a stereomicroscope. Therefore, experience in operating under a microscope and perfecting a successful craniotomy is essential. It is best to establi...

<table border="1">
 	<tbody>
 	<tr>
 	<th>NAME</th>
 <th>COMPANY AND CATALOGUE NUMBER</th>
 </tr>
 <tr>
 	<td>A. Solutions and Chemicals</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 	<td>Povidone-Iodine Solution, Surgical Scrub </td>
 <td>Aplicare, 82-209</td>
 </tr>
 <tr>
 	<td>70% Ethanol</td>
 <td>Koptec, V1001</td>
 </tr>
 <tr>
 	<td>2,3,5 Triphenyltetrazolium chloride (TTC)</td>
 <td>Sigma, T8877</td>
 </tr>
 <tr>
 	<td>4% Paraformaldehyde </td>
 <td>Electron Microscopy Sciences, 159SP</td>
 </tr>
 <tr>
 	<td>Phosphate Buffered Saline, pH:7.4</td>
 <td>Made and sterilized in the lab.</td>
 </tr>
 <tr>
 	<td>Mineral Oil </td>
 <td>PML Microbiologicals, R6570</td>
 </tr>
 <tr>
 	<td>Isofluorane</td>
 <td>The Butler Company, 029405</td>
 </tr>
 <tr>
 	<td>Buprenorphine (0.3mg/ml)</td>
 <td>Hospira, NDC 0409-2012-32</td>
 </tr>
 <tr>
 	<td>Artificial tears</td>
 <td>The Butler Company, 007312</td>
 </tr>
 <tr>
 	<td>Recovery Gel</td>
 <td>Clear H2O, 72-01-5022</td>
 </tr>
 <tr>
 	<td>B. Surgical Materials and Equipment</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 	<td>C57/BL6 mouse, 10-12 weeks old</td>
 <td>The colony was bred in the university vivarium</td>
 </tr>
 <tr>
 	<td>100% oxygen tank</td>
 <td>Airgas</td>
 </tr>
 <tr>
 	<td>Anesthetic Vaporizer and Flow meter</td>
 <td>Surgivet, Model 100</td>
 </tr>
 <tr>
 	<td>Heating Pad with Rectal Probe</td>
 <td>Fine Science Tools, TR200</td>
 </tr>
 <tr>
 	<td>Dissecting microscope</td>
 <td>Zeiss, Stemi 2000</td>
 </tr>
 <tr>
 	<td>Heating panel</td>
 <td>Petco Services Ltd. 0307-013</td>
 </tr>
 <tr>
 	<td>Small Vessel Cauterizer Set</td>
 <td>Fine Science Tools, 18000-00</td>
 </tr>
 <tr>
 	<td>Mini Peristaltic Pump</td>
 <td>Harvard Apparatus, MPII</td>
 </tr>
 <tr>
 	<td>1ml syringe</td>
 <td>BD, 329650</td>
 </tr>
 <tr>
 	<td>18G needle</td>
 <td>Becton Dickinson, 305196</td>
 </tr>
 <tr>
 	<td>Surgical 5.0 Nylon Suture</td>
 <td>Ethilon, 698H</td>
 </tr>
 <tr>
 	<td>Sectioning Block</td>
 <td>Kent Scientific, 1mm</td>
 </tr>
 <tr>
 	<td>Single Edge Razor Blades</td>
 <td>Electron Microscopy Sciences, 71962</td>
 </tr>
 <tr>
 	<td>Label Tape</td>
 <td>Scienceware, Bel-art Products F13463-2075</td>
 </tr>
 <tr>
 	<td>Gauze Sponges</td>
 <td>Kendall Curity 2252, Johnson and Johnson 6415</td>
 </tr>
 <tr>
 	<td>Cotton Tip Applicators</td>
 <td>Puritan 806-WCL</td>
 </tr>
 <tr>
 	<td>C. Surgery Tools</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 	<td>Surgical Spring Scissors</td>
 <td>Fine Science Tools 15003-08</td>
 </tr>
 <tr>
 	<td>Microdissecting forceps angled</td>
 <td>Fine Science Tools 5/45</td>
 </tr>
 <tr>
 	<td>Microdissecting forceps curved</td>
 <td>Fine Science Tools 5 Ti</td>
 </tr>
 <tr>
 	<td>Fine straight scissors</td>
 <td>Fine Science Tools 14568-12</td>
 </tr>
 <tr>
 	<td>Curved bone rongeur</td>
 <td>Fine Science Tools 16021-14</td>
 </tr>
 <tr>
 	<td>Hartman Hemostat Curved</td>
 <td>Fine Science Tools 13003-10</td>
 </tr>
 </tbody>
</table>

Table 1. The list of materials, tools and chemicals used for the MCA ligation procedure. A. Solutions and Chemicals, B. Surgical Materials and Equipment, C. Surgery Tools.

the application of permanent middle cerebral artery ligation in the mouse

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to develop suitable animal models to study the events that unfold during ischemic insult. These techniques include transient or permanent, focal or global ischemia models using many different animal models, with the most common being rodents.
The permanent MCA ligation method which is also referred as pMCAo in the literature is used extensively as a focal ischemia model in rodents 1-6. This method was originally described for rats by Tamura et al. in 1981 7. In this protocol a craniotomy was used to access the MCA and the proximal regions were occluded by electrocoagulation. The infarcts involve mostly cortical and sometimes striatal regions depending on the location of the occlusion. This technique is now well established and used in many laboratories 8-13. Early use of this technique led to the definition and description of &#x201C;infarct core&#x201D; and &#x201C;penumbra&#x201D; 14-16, and it is often used to evaluate potential neuroprotective compounds 10, 12, 13, 17. Although the initial studies were performed in rats, permanent MCA ligation has been used successfully in mice with slight modifications 18-20 .
This model yields reproducible infarcts and increased post-survival rates. Approximately 80% of the ischemic strokes in humans happen in the MCA area 21 and thus this is a very relevant model for stroke studies. Currently, there is a paucity of effective treatments available to stroke patients, and thus there is a need for good models to test potential pharmacological compounds and evaluate physiological outcomes. This method can also be used for studying intracellular hypoxia response mechanisms in vivo. 
Here, we present the MCA ligation surgery in a C57/BL6 mouse. We describe the pre-surgical preparation, MCA ligation surgery and 2,3,5 Triphenyltetrazolium chloride (TTC) staining for quantification of infarct volumes.

Watch this Scientific Journal Video about The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse at JoVE.com

The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse

Middle cerebral artery (MCA) ligation is a technique to study focal cerebral ischemia in animal models. In this method, the middle cerebral artery is exposed by craniotomy and ligated by cauterization. This method gives highly reproducible infarct volumes and increased post-operative survival rates compared to other methods available.

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to ...

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